The overall aims of this project are to compare aerosol administration, tracheal instillation, and bronchial artery injection as methods of delivery for adenoviral vectors and other vectors to the airway ofanimals in preparation for human clinical trials. Clinical trials in CF patients are to be developed as part of this project, but specific protocols are not available and approved at this time. An adenoviral (Ad) vector expressing beta-galactosidase currently available from Dr. Graham will be used to assess aerosol delivery to the airway of first rats and then baboons in comparison to tracheal instillation focusing particularly on the distribution and proportion of cells expressing beta-galactosidase at various levels of the respiratory tract and quantitating the amount of inflammation produced. Ad vectors expressing beta-galactosidase will be used to assess delivery to the airway by injection into the bronchial artery of sheep, and this will be compared to the results with aerosol administration and tracheal instillation; bronchial artery injection will be extended to baboons if promising in sheep. An Ad vector expressing CFTR developed in collaboration with Dr. Graham will be used for preclinical studies in baboons and cotton rats to establish baseline information for development of human clinical trials. Based on the above results, human clinical trials will be developed and will likely focus on the use of aerosol administration or bronchial artery injection if either of these show any advantages over tracheal instillation. The human work will begin immediately with assessment of the natural history of adenovirus infections CF patients and with in vivo electrophysiological measurements in patients. New Ad vectors developed in the Graham project, nonviral vectors developed in the Smith project, and AAV vectors provided by collaborators will be tested for airway delivery first in the rat using a nuclear targeted beta-galactosidase reporter gene or other reporter gene to measure toxicity, percent of transfected cells, level of airway delivery, and duration of expression. Any new vectors demonstrating potentially promising results in the rat will then be studied in baboons, and will be brought to human clinical trials as warranted by the results. CF mutant mice will be evaluated for pulmonary pathology using strategies to prolong survival and using infection with respiratory syncytial virus, Staph. aureus, and Pseudomonas in collaborative studies. CF mutant mice will be used to test the ability to correct the electrophysiological defect and any pathological abnormalities in the lung using Ad vectors expressing CFTR.